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Validated biomarkers predictive of response/resistance to anthracyclines in breast cancer are currently lacking. The neoadjuvant Trial of Principle (TOP) study, in which patients with estrogen receptor (ER)–negative tumors were treated with anthracycline (epirubicin) monotherapy, was specifically designed to evaluate the predictive value of topoisomerase II-alpha (TOP2A) and develop a gene expression signature to identify those patients who do not benefit from anthracyclines. Here we describe in details the contents and quality controls for the gene expression and clinical data associated with the study published by Desmedt and colleagues in the Journal of Clinical Oncology in 2011 (Desmedt et al., 2011). We also provide R code to easily access the data and perform the quality controls and basic analyses relevant to this dataset.

The pathogenesis of breast cancers that do not express estrogen receptors or Her-2/neureceptors (ER−/HER2− phenotype) is incompletely understood. We had observed markedly elevated gene expression of gamma-aminobutyric acid type A (GABAA) receptor subunit π (GABAπ, GABRP) in some breast cancers with ER−/HER2− phenotype. In this study, transcriptional profiles (TxPs) were obtained from 82 primary invasive breast cancers by oligonucleotide microarrays. Real-time reverse transcription–polymerase chain reaction (RT–PCR) was used to measure GABAπ gene expression in a separate cohort of 121 invasive breast cancers. GABAπ gene expression values from TxP and RT–PCR were standardized and compared with clinicopathologic characteristics in the 203 patients. GABAπ gene expression was increased in 16% of breast cancers (13/82 TxP, 20/ 121 RT–PCR), particularly in breast cancers with ER−/HER2− phenotype (60%), and breast cancers with basal-like genomic profile (60%). The profile of genes coexpressed with GABAπ in these tumors was consistent with an immature cell type. In multivariate linear regression analysis, the level of GABAπ gene expression was associated with ER−/HER2− phenotype (P<0.0001), younger age at diagnosis (P=0.0003), and shorter lifetime duration of breastfeeding (≤ 6 months) in all women (P=0.017) and specifically in parous women (P=0.013). GABAπ gene expression was also associated with combinations of high grade with ER−/HER2− phenotype (P=0.002), and with Hispanic ethnicity (P=0.036). GABAπ gene expression is increased in breast cancers of immature (undifferentiated) cell type and is significantly associated with shorter lifetime history of breastfeeding and with high-grade breast cancer in Hispanic women.

Microtubule binding protein Tau was recently identified through gene expression analysis of human breast cancer tissues as a novel marker of response to paclitaxel.1 This article reviews these recent findings and provides additional information to support the role of Tau as an emerging marker and mediator of paclitaxel sensitivity. Low expression of Tau is associated with increased sensitivity to paclitaxel in human breast cancer as well as in a broad range of cell lines. Down regulation of Tau in cell lines by siRNA increases their sensitivity to paclitaxel but not to anthracycline chemotherapy. We propose that this is due to increased paclitaxel binding to microtubules when microtubules are assembled in the presence of low concentrations (or absence) of Tau compared to microtubules that are formed in the presence of physiological (or higher) levels of Tau.

Individualized selection of chemotherapy regimens based on the molecular characteristics of the cancer would be desirable. However, it is not currently feasible. The development of clinically useful predictors of response to chemotherapy has proven to be difficult. Recently, novel analytical tools particularly transcriptional profiling, have shown promise as potential predictive tests in several small exploratory studies. However, many of the practical challenges of clinical pharmacogenomics are the same that have plagued prognostic and predictive marker research for decades. Perhaps, the most important challenge is to prospectively design and conduct validation trials that demonstrate clinical utility by showing improved patient outcomes with the use of a proposed new test.

The sigma 1 receptor (S1R) represents a unique drug-binding site that is distinct from any other receptors. We examined S1R expression in human breast cancer and assessed the activity of S1R ligands in breast cancer cell lines. One-hundred nine breast specimens from normal breast, benign breast disease and cancer were examined with immunohistochemistry or RT- PCR and six different cell lines were also evaluated. S1R mRNA overexpression was detected in 64% of breast cancers compared to normal breast tissue. Immunohistochemistry showed positive epithelial cell staining in 60% of invasive and 41% of in situcancers, 75% of ductal hyperplasia and in 33% of normal breast. The pattern of expression was more diffuse in invasive breast carcinoma compared to other conditions (p= 0.02). S1R expression was neither a prognostic nor a predictive factor for efficacy of adjuvant chemotherapy but the study only included 58 cancer patients and therefore the statistical power is limited. MDA - MB −361, MDA - MB −435, BT 20 and MCF 7 cells all expressed S1R protein by Western blot. The non-specific S1R ligands haloperidol, reduced haloperidol and progesterone produced a dose-dependent inhibition of the growth at high (>10 μM) concentrations. Reduced haloperidol also showed additive cytotoxic effects when combined with doxorubicin, vinorelbine , paclitaxel and docetaxel in vitro. The S1R-specific ligand, SKF 10047 demonstrated the least growth inhibitory activity and showed no interaction with chemotherapy. These results demonstrate that some normal and most neoplastic breast epithelial cells and cell lines commonly express S1R. High concentrations of haloperidol inhibit the growth of these cells and potentiate the effect of chemotherapy in vitro.

The aim was to assess chemotherapy-induced histologic changes in breast cancer and to evaluate their relationship with tumour response, to treatment and patient survival. We examined paired pre- and post-chemotherapy tumour specimens from 57 patients with stages II and III breast cancers. All patients received three to four courses of anthracycline-containing preoperative treatment and subsequently underwent modified radical mastectomy 3–5 months after diagnosis. Histologic parameters evaluated included extent of residual disease and cytologic changes in residual tumour and in non-neoplastic breast tissue and lymph nodes. Correlation of pathologic changes with disease-free and overall survival was evaluated. Clinical evaluation indicated that 24% (n=14) of patients had a complete response, 53% (n=30) had a partial response, and 22% (n=13) had stable disease. Pathologic assessment revealed a complete response in 14% (n=8) of patients. Clinical and histologic complete responses were discordant in 35% of patients. Sixty-eight percent of tumours demonstrated moderate to severe cytologic changes in the cytoplasm and nucleus of neoplastic cells. These were characterized by increased overall cell size, cytoplasmic vacuolization, nuclear enlargement, multinucleation, and vesiculation of chromatin. Chemotherapy-induced histologic changes varied within a tumour and between patients. Tumour cytologic changes were significantly associated with histologic (p=0.03) as well as clinical (p=0.02) tumour response. Pathologic tumour response, but not histologic changes, was also a powerful predictor of overall survival and relapse-free survival. In the non-neoplastic glandular tissue, periductal and perilobular fibrosis and atrophy of lobular acini similar to those seen in postmenopausal breast tissue were noted. Changes in lymph nodes were characterized by lymphoid depletion and fibrosis with and without tumour metastasis. Chemotherapy-induced cytologic changes are frequently seen in neoplastic breast epithelium and correlate with response to treatment. Histologic confirmation of clinical tumour response to preoperative chemotherapy is necessary for accurate characterization of patients', response to treatment.

MCGR (Monte Carlo determination of G(r)) is an inverse method for determining the pair correlation function from the structure factor. In this paper we describe the algorithm in detail, including recent developments such as the possible application of coordination constraints, Gaussian smoothing of peak shapes and the ability to subtract a (quadratic) polynomial background. We will illustrate how these have important practical implications for the way that experiments can be performed. Examples of the applications of MCGR to liquids/glasses and crystalline materials will be given.

Recent advances in understanding how response or resistance to cytotoxic drugs develops at the cellular level resulted in the development of novel, non-cytotoxic agents that modulate response the chemotherapy. ‘Chemo-signal therapy’, the combination of chemotherapy with cellular response modifiers, is a very promising new treatment modality that has entered the arena of clinical trials. Clinical experience with the anti-HER-2 antibody, trastuzumab, in breast cancer has demonstrated that manipulation of growth factor signalling can enhance sensitivity to cytotoxic drugs in a clinically meaningful way. Several other agents that were designed to modulate response to chemotherapy are currently in early phases of clinical drug development. It is likely that some of these new molecules will have a major impact on how chemotherapy will be given in the next decade. This paper will review current clinical research with a select group of chemotherapy response modifiers. We will focus on agents that modulate signal transduction, oncogene expression and apoptosis with an emphasis on breast cancer.

MCGR (Monte Carlo determination of G(r)) is an inverse method for determining the pair correlation function from the structure factor. In this paper we describe the algorithm in detail, including recent developments such as the possible application of coordination constraints, Gaussian smoothing of peak shapes and the ability to subtract a (quadratic) polynomial background. We will illustrate how these have important practical implications for the way that experiments can be performed. Examples of the applications of MCGR to liquids/glasses and crystalline materials will be given.

A simple numerical method is proposed whereby the relative information content of different total structure factors can be determined, whether obtained by neutron diffraction with isotopic substitution, X-ray diffraction with anomalous scattering or EXAFS. By taking into account the likely level of experimental errors it can then be decided which experiments provide the maximum information and which provide no additional information. The method is illustrated by creating different sets of data corresponding to a two component Lennard Jones liquid, and then determining the partial radial distribution functions From these sets of data.

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A simple numerical method is proposed whereby the relative information content of different total structure factors can be determined, whether obtained by neutron diffraction with isotopic substitution, X-ray diffraction with anomalous scattering or EXAFS. By taking into account the likely level of experimental errors it can then be decided which experiments provide the maximum information and which provide no additional information. The method is illustrated by creating different sets of data corresponding to a two component Lennard Jones liquid, and then determining the partial radial distribution functions from these sets of data.

This report reviews clinical experience with high-dose chemotherapy with stem-cell support in breast cancer and attempts to integrate clinical and molecular biological observations into a model of drug resistance. Despite the remarkable initial chemosensitivity of breast cancer, the majority of patients with advanced breast cancer die of their disease. In trials to date, high-dose chemotherapy has not been able to overcome primary drug resistance and patients with disease progression during induction therapy benefit little from further high-dose therapy. On the other hand, high-dose chemotherapy produces improved objective response rates compared with standard-dose chemotherapy due to converting some partial responses achieved by standard-dose induction therapy to complete response. This suggests that high-dose chemotherapy may overcome partial clinical drug resistance and may result in a more complete elimination of chemosensitive cells. Whether increased complete response rates will translate into higher cure rates and increased overall survival remains to be unequivocally demonstrated. There are multiple clinical patterns of drug failure including continued growth during chemotherapy, partial response followed by a period of stable disease, initial complete response with subsequent recurrence within a few months or after several years. Different mechanisms of drug resistance may operate at different stages of the disease and predispose patients to different clinical patterns of failure. A model of clonal progression of cancer is proposed that could explain several intriguing features of clinical drug resistance. We hypothesize that drug-sensitivity is an acquired characteristic of neoplastic cells and that a 'physiological drug-resistant' state may precede drug sensitivity at early stages of neoplastic transformation. Some recurrences may, in this context, represent progression of physiologically drug-resistant clones to sensitivity and subsequently to 'pathologic resistance' to chemotherapy. Optimal therapy to delay or prevent recurrence may differ depending on the stage and biology of the tumor and may include combinations of cytotoxic drugs and chemopreventive agents to arrest progression of early physiologically drug-resistant neoplastic stem cells.

The structures of 51 elemental liquids have been modelled from experimental diffraction data using the reverse Monte Carlo method. The structural trends across the periodic table are described in terms of three-body correlations and discussed in terms of the relative importance of two-body and many-body forces.

A method has been developed for the study of vacancy dynamics in molecular dynamics simulations of either liquids or solids. This has been applied to the case of molten RbCl to show that vacancies may be considered to exist in the liquid state using either static or dynamic criteria, but that these do not represent a significant proportion of the free volume.

The expression of tumour necrosis factor alpha (TNF-alpha) and its two distinct receptors, TNF-R p55 and TNF-R p75, was assessed by immunocytochemistry in 28 primary breast cancer and three reduction mammoplasty specimens ('normal' breast tissue). Expression of TNF-alpha or TNF-R p75 was not detectable in normal breast tissue or in non-malignant breast tissue adjacent to the tumours. By contrast, TNF-R p55 was expressed by occasional stromal cells in normal tissue. TNF-alpha was expressed focally in 50% of the tumours studied, being largely localised to macrophage-like cells in the stroma. TNF-R p55 was expressed by a population of stromal cells in all the tumours examined, and a varying proportion of neoplastic cells in 75% of these tissues. TNF-R p75 was detected in about 70% of the tumours, immunoreactivity being confined mainly to cells in the stroma. In this preliminary study there was no association between the above cytokine parameters and such measures of tumour biology as lymph node status, tumour grade, proliferative activity or degree of angiogenesis. However, there was a correlation between the expression of TNF-R p55 by blood vessels and the number of leucocytes present.

The effects of tumour necrosis factor-alpha (TNF alpha) on the growth and DNA synthesis of the human breast cell line, T47D, were studied. A dose-dependent, reversible inhibition of thymidine incorporation and cell growth was observed in the range of 0.1 ng ml-1 to 100 ng ml-1 of TNF alpha. Cell viability was not impaired in any of the experiments. Flow-cytometric DNA analysis demonstrated that after 24 h exposure to TNF alpha, T47D cells accumulated in the G1 phase of the cell cycle, and were depleted in the G2/M and S phases, suggesting a block in the progression of the G1/S transition. The involvement of protein kinases (PK) and protein phosphatases in TNF alpha-induced signal transduction was also investigated. A transient and rapid 2-fold increase in total cellular protein kinase C (PKC) activity was detected after 10 min exposure to TNF alpha. To study the role of the observed PKC activation in the cytostatic effect of TNF alpha, T47D cells were exposed to the cytokine in the presence of the potent PKC inhibitor, H7. The inhibitory effect of TNF alpha on thymidine incorporation was not affected by exposure to H7 at concentrations sufficient to block the stimulation of thymidine up-take induced by the PKC agonist, phorbol-12-myristate-13-acetate (PMA). The involvement of other signalling pathways was addressed using the cyclic nucleotide-dependent PK inhibitor, H8; the calmodulin-dependent PK inhibitor, W7; and the inhibitor of protein phosphatases PP1 and PP2B, okadaic acid. Exposure of T47D cells to these enzyme inhibitors failed to antagonise the inhibition of thymidine incorporation by TNF alpha. Taken together, these results indicate that the cytostatic effect of TNF alpha on T47D cells occurs at the G1/S transition of the cell cycle, and is mediated by an intracellular pathway which does not involve the activity of protein kinases C and A, nor protein phosphatases PP1, PP2B.

Relationship of corticotroph proliferation answer and survival time after adrenalectomy was examined. Corticotroph proliferation rates were detected by short-term 3H-thymidine radiolabeling, then ACTH immunostaining and autoradiography. Effect of adrenalectomy on corticotroph proliferation rate was examined in vivo and an elevation was demonstrated first on the second postoperative day, increasing on the third-fourth day postoperatively and then decreasing. Effects of different secretagogues on corticotroph proliferation were examined in short-term pituitary monolayer cultures taken from ADX rats. CRF and Forskolin treatment potentiated corticotroph proliferation in cultures taken from adrenalectomized rats, but not in the controls. We suggest that croticotroph proliferation is stimulated via the cAMP-proteinkinase A pathway, while adrenalectomy plays a permissive role.

Extensive Reverse Monte Carlo (RMC) model calculations have been performed for several hydrogenated amorphous carbon systems, for which only one, or at most two, neutron diffraction measurement(s) had been carried out. The possibility of determining the microscopic density of the samples, of estimating the chemical composition of the materials, and of deriving reliable (partial) pair correlation functions from reduced-range (Qmax< 15 Å−1) structure factors have been investigated in particular. The number density could be determined within 10% in most of the (model) cases, whereas the estimation of the composition proved to be successful only in one of the four cases studied here. It is shown that an evaluation of the partial pair correlation functions (ppcf) for these materials is possible on the basis of limited scattering vector range.

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We have developed a new technique, based on the standard Monte Carlo simulation method with Markov chain sampling, in which a set of three dimensional particle configurations are generated that are consistent with the experimentally measured structure factor. A(Q), and radial distribution function, g(r), of a liquid or other disordered system. Consistency is determined by a standard χ2 test using the experimental errors. No input potential is required, we present initial results for liquid argon. Since the technique can work directly from the structure factor it promises to be useful for modelling the structures of glasses or amorphous materials. It also has other advantages in multicomponent systems and as a tool for experimental data analysis.

Background: Both hypothermia and CNS drugs may alter the prognostic accuracy of aEEG in HIE. We aimed to assess the predictive value of aEEG in hypothermia treated HIE infants. Furthermore we intended to investigate the association of cumulative doses of CNS drugs with aEEG recovery.Patients and methods: 63 term HIE infants treated with moderate systemic hypothermia for 72 hours were continuously monitored by single-channel aEEG. Records were evaluated according to the classification suggested by Hellström-Westas (2006., Seminars in Fetal & Neonatal Medicine). Doses of administered Morphine, Phenobarbitone and Midazolam were recorded. Poor outcome was defined as death or severe neurodevelopmental delay at 18-22 months (Bayley Scales of Infant Development II), good outcome as absence of these criteria. Mann-Whitney U test was used for statistical analysis.Results: Poor outcome n= 25, good outcome n= 38. PPVs of an abnormal background pattern to predict poor outcome were 0.5 at 6hs; 0.7 at 24hs; 0.88 at 48hs and 0.95 at 60hs. Cumulative doses of the investigated drugs did not differ between infants with onset of normal background pattern before and after 24 hours.Conclusions: aEEG provides the best prediction of outcome from the 48th hour during hypothermia in HIE infants. Commonly used CNS drugs in HIE infants do not seem to considerably delay aEEG recovery.Figure